Comparison of the neural differentiation potential of human mesenchymal stem cells from amniotic fluid and adult bone marrow.

Human mesenchymal stem cells (MSCs) are considered a promising tool for cell-based therapies of nervous system diseases. Bone marrow (BM) has been the traditional source of MSCs (BM-MSCs). However, there are some limitations for their clinical use, such as the decline in cell number and differentiation potential with age. Recently, amniotic fluid (AF)-derived MSCs (AF-MSCs) have been shown to express embryonic and adult stem cell markers, and can differentiate into cells of all three germ layers. In this study, we isolated AF-MSCs from second-trimester AF by limiting dilution and compared their proliferative capacity, multipotency, neural differentiation ability, and secretion of neurotrophins to those of BM-MSCs. AF-MSCs showed a higher proliferative capacity and more rapidly formed and expanded neurospheres compared to those of BM-MSCs. Both immunocytochemical and quantitative real-time PCR analyses demonstrated that AF-MSCs showed higher expression of neural stemness markers than those of BM-MSCs following neural stem cell (NSC) differentiation. Furthermore, the levels of brain-derived growth factor and nerve growth factor secreted by AF-MSCs in the culture medium were higher than those of BM-MSCs. In addition, AF-MSCs maintained a normal karyotype in long-term cultures after NSC differentiation and were not tumorigenic in vivo. Our findings suggest that AF-MSCs are a promising and safe alternative to BM-MSCs for therapy of nervous system diseases.

Vasculogenic mimicry is a prognostic factor for postoperative survival in patients with glioblastoma.

A previous report has confirmed the existence and clinical significance of vasculogenic mimicry (VM) in glioma. However, its conclusions about the negative clinical significance of VM in glioblastoma are based on a small group of patients and, thus, might be unconvincing. The aim of the present study was to reevaluate the clinical significance of VM in glioblastoma. Patients were classified as VM-positive or VM-negative according to CD34 and periodic acid-Schiff staining. The association between VM and the clinical characteristics of the patients was analyzed. Univariate and multivariate analyses were carried out to identify the independent prognostic factors for overall survival using the Cox regression hazard model. Survival times were estimated using the Kaplan-Meier method and compared using the log-rank test. Of all 86 glioblastomas, 23 were found to have VM. The presence of VM in glioblastoma was not associated with gender, age, Karnofsky performance status, hydrocephalus, tumor burden, microvessel density, tumor relapse, or the extent of tumor resection. The univariate and multivariate analyses revealed that VM is an independent prognostic factor for overall survival. The median survival time for patients with VM was 11.17 months compared with 16.10 months for those without VM (P = 0.017). In addition to VM, an age of 65 years or older, a KPS of 60 or less, a large tumor burden are significant prognostic factors for patient survival. Our data suggest that VM might be an independent adverse prognostic factor in newly diagnosed GBM, further prospective studies are needed to answer this question.

Vasculogenic mimicry and its clinical significance in medulloblastoma.

Vasculogenic mimicry (VM), a process involving the formation of a tubular structure by highly invasive and genetically dysregulated tumor cells, can supplement the function of blood vessels to transport nutrients and oxygen to maintain the growth of tumor cells in many malignant tumors. We aimed to explore the existence of VM and its clinical significance in medulloblastoma in this study. VM was identified in 9 out of 41 (22%) medulloblastoma tissues. Immunohistochemical studies revealed that the presence of VM was associated with the expression of MMP-2, MMP-14, EphA2 and laminin 5γ2. Tumor tissues with VM were associated with lower microvessel density (MVD), which was indirect evidence of the blood supply function of VM. Survival analysis and log-rank tests showed that patients with VM had shorter overall survival time than those without VM. Multivariate analysis and the Cox proportional hazards model identified VM as independent prognostic factor for overall survival. Our results confirmed the existence of VM for the first time and revealed that VM is a strong independent prognostic factor for survival in patients with medulloblastoma.

Molecular targeting of malignant glioma cells with an EphA2-specific immunotoxin delivered by human bone marrow-derived mesenchymal stem cells.

Immunotoxins have shown great promise as an alternative treatment for brain malignancies such as gliomas, but their failure to penetrate into the tumor mass remains a major problem. Mesenchymal stem cells exhibit tropism to tumor tissue and may serve as a cellular vehicle for the delivery and local production of antitumor agents. In this study, we used human bone marrow-derived mesenchymal stem cells (hMSCs) as a vehicle for the targeted delivery of EphrinA1-PE38, a very specific immunotoxin against the EphA2 receptor that is overexpressed in gliomas. hMSCs were transduced with adenovirus to express secretable EphrinA1-PE38. Our invitro assays confirmed the expression, release and selective killing effect of the immunotoxin produced by hMSCs. Furthermore, the intratumoral injection of engineered hMSCs was effective at inhibiting tumor growth in a malignant glioma tumor model. These results indicate that gene therapy utilizing EphrinA1-PE38-secreting hMSCs may provide a novel approach for the local treatment of malignant gliomas.

Investigation of a plasmid containing a novel immunotoxin VEGF165-PE38 gene for antiangiogenic therapy in a malignant glioma model.

Inhibition of tumor neovascularization has profound effects on the growth of solid tumors. Our previous studies have shown the effect of VEGF165-PE38 recombinant immunotoxin on proliferation and apoptosis in human umbilical vein endothelial cells in vitro. In this study, we explored the direct inhibition of angiogenesis in chick chorioallantoic membrane and antiangiogenic therapy in a malignant glioma model. HEK293 cells were transfected with the pVEGF165PE38-IRES2-EGFP plasmid. ELISA was used to confirm the expression of VEGF165-PE38 in the transfected cells. These cells released 1396 + or - 131.9 pg VEGF165-PE38/1x10(4) cells/48 h into the culture medium and the supernatant was capable of inhibiting the growth of capillary-like structures in chick chorioallantoic membrane assay. In a murine malignant glioma model, plasmid was directly administered via multiple local intratumoral delivery. After day 16 the tumor volume in mice treated with pVEGF165PE38-IRES2-EGFP was significantly lower than that in mice in the control groups. Immunohistochemistry studies showed that the treated group had decreased expression of CD31. Quantitative analysis of microvessel density in the treated group was 1.99 + or - 0.69/0.74 mm(2), and was significantly lower than that in the control groups (9.33 + or - 1.99/0.74 mm(2), 8.09 + or - 1.39/0.74 mm(2) and 8.49 + or - 1.69/0.74 mm(2)). Immunohistochemistry analysis indicated that immunotoxin VEGF165-PE38 was distributed in the treated group in malignant glioma tissue. Our findings provide evidence that the in vivo production of VEGF165-PE38 through gene therapy using a eukaryotic expression plasmid had potential antiangiogenic activity in malignant glioma in vivo.

[Preparation and property of platinum microcoil modified by a copolymer-VEGF conjugate].

OBJECTIVE: To prepare a platinum microcoil coated with polymers and vascular endothelial growth factor (VEGF), and evaluate its surface characteristics and property of sustained VEGF release. METHODS: The surface of the platinum microcoils (GDC) were modified by coating P(DLLA-co-TMC) copolymer and immobilizing heparin on the surface of GDC. VEGF was then loaded onto the surface of GDC and the controlled release of VEGF within GDC was achieved. The morphology was observed by scanning electron microscope, and the sustained release of VEGF was evaluated by enzyme-linked immunosorbent assay (ELISA). RESULTS: Platinum coils were prepared by successive deposition of P(DLLA-co-TMC) copolymer and anionic heparin, and VEGF was immobilized through affinity interaction with heparin. The accumulative release of VEGF increased obviously during the entire testing period without burst release. CONCLUSION: The use of P(DLLA-co-TMC) copolymer allows immobilization of VEGF on the platinum coils for controlled VEGF release, and improves the biological property of the coils.

Human mesenchymal stem cells-like cells as cellular vehicles for delivery of immunotoxin in vitro.

Human mesenchymal stem cells-like cells (hMSCs-like cells) were used as a tumor treatment platform for the systemic delivery of immunotoxin genes. VEGF165-PE38 recombinant immunotoxin served as the model system. hMSCs-like cells were isolated, expanded, and electroporated with the pIRES2-VEGF165PE38-EGFP plasmid. RT-PCR and ELISA were used to confirm the expression of VEGF165-PE38 in the transfected hMSCs-like cells. These cells released 1390 +/- 137 pg VEGF165-PE38/10(4)cells over 48 h into the culture medium and the supernatant was capable of selectively killing human umbilical vein endothelial cells (HUVECs) and increasing apoptosis in these cells. In contrast, RPMI8226 was not inhibited by identical supernatants. Thus, these results lay the foundation for further studies on the potential role of hMSCs-like cells as a targeted therapeutic delivery vehicle for immunotoxins.